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1. λ lysogens are immune to superinfection by λ, but not with other lambdoid phages such as 434. λimm434 (a lambdoid hybrid phage that has mostly λ genes except for cI and the surrounding “immunity” region of 434 phage) also can grow on E. coli(λ). a. If you take a 1ml culture of E. coli(λ) (this is wild-type E. coli) and superinfect with λimm434 at an MOI of 1, the tube will clear after about an hour. Describe how you would determine whether the released phages from this infection are λimm434 or λ (induced from the prophage). Do you expect there will be more λimm434, more λ, or equal numbers of both phages in the lysate? Way more λimm434. b. You perform infection experiments using wild-type E. coli and E. coli(λ), infecting them at an MOI = 1 with either λ, 434, λimm434, or λimm434 Ram (an amber mutant in the R gene) and get the following result: λ E. coli + E. coli(λ) + = lysis, - = no lysis Infecting Phage 434 λimm434 + + + + λimm434 Ram + Explain, in detail, why λimm434 Ram is able to lyse the λ lysogen but not wild-type E. coli. It doesn’t lyse wild-type because it is deficient in R, one of the two lysin genes. The reason why infection of the lysogen leads to lysis is because the R gene on the prophage is transcriptionally activated and translated, complementing the mutant. The reason why R is transcribed, as well as all of the late genes, is because Q is supplied in trans by λimm434. The important thing here is that while PR and PL are directly repressed by cI, PR’ is not and RNAP able to bind and initiate transcription there, even in the prophage state. c) Do you expect there are viable phages in the lysate from the infection of E. coli(λ) with λimm434 Ram and, if so, what phage genotype will be present? λimm434 Ram